A Novel LC-MS/MS Method for Improving Long-Term Stability of Retinol in Dried Blood Spots and Quantification of Its Levels

Abstract

Abstract Objectives To establish an accurate and reliable method for stabilizing vitamin A (retinol) in Dried blood spots (DBS), quantifying and comparing DBS retinol concentrations with their equivalent plasma retinol levels. Methods Antioxidants pretreated on paper combined with vacuum treatment were used to increase retinol stability on DBS. A surrogate matrix of whole blood prepared using mixture of human erythrocytes and 2% BSA in PBS was firstly used in DBS retinol determination based on the fact that retinol is excluded from erythrocytes. Results DBS retinol was stable during 120 min of air drying and 30 days of room-temperature storage. The method was linear in the concentration range of 0.04–300 μg/mL. Accuracy was calibrated using two National Institute of Standards and Technology (NIST) calibrants generated serum at concentrations of 0.1962 and 0.3948 g/mL, relative errors (RE% values) of 0.07% and 4.95% were found, respectively. Both the between-run (n = 5) and within-run (n = 6) precision (relative standard deviations, RSD%) were below 8.42%. The spiked recoveries at 3 concentrations ranged from 86.48% to 98.13%. A reliable calibration model was first developed to convert DBS retinol concentration to the equivalent plasma retinol concentration. Conclusions The validated method can be applied to the nutritional assessment of vitamin A, using the established calibration model, DBS retinol can compare with clinical reference ranges and with studies using serum or plasma samples. Funding Sources The National Key Research and Development Program of China (2018YFC1002503); The National Natural Science Foundation of China (Grant No. 81,400,848, 81,701,441); The CAMS Initiative for Innovative Medicine (2016-I2M-1–008); The Beijing municipal program of medical research (Grant No. 2016–04); The National Key Research and Development Program of China (No. 2016YFC1306204).